There is an urgent need for clinically useful point of care (POC) technologies for pathogen identification, particularly in nontraditional healthcare settings. The goal of this collaborative research proposal is to develop a highly portable diagnostic platform for POC diagnosis of urinary tract infections (UTI). The core of this platform, Portable Pathogen Alert Laboratory (P-PAL), is a well-established, clinically validated electrochemical biosensor array integrated with microfluidic technology combined with innovative strategies to expedite sample processing using electrokinetic forces. We have assembled an academic-industry partnership with complementary expertise in electrochemical biosensors (V.G.), microfluidics/electrokinetics (P.W.), urology (J.L.), clinical microbiology (E.J.B.), and biostatistics (M.S.). Our team previously described the electrochemical biosensor for rapid identification of uropathogens in clinical urine samples. The assay involves sandwich hybridization of bacterial target 16S rRNA to species-specific capture and detector probes. At present, the assay is not practical for POC application given the requirement for manual sampleprocessing steps including centrifugation, mixing and reagent transferring. Additional technology development and system integration are needed to enhance the sensitivity, speed and to enable POC utility. We propose 4 Specific Aims: (1) To develop electrokinetic and microfluidic sample processing modules using a multi-functional 'universal electrode'; (2) To develop the P-PAL Cartridge composed of the universal electrode array, the sample processing fluidic handling cartridge and a reagent storage cartridge; (3) To integrate all the fluidic, pneumatic, and electrical controls for the P-PAL System for reagent delivery, reagent-sample mixing, and target detection; and (4) To validate P-PAL using spiked urine samples and unknown clinical urine samples. We anticipate that we will achieve molecular pathogen identification within 15 minutes with detection sensitivity of 1000 cfu/ml. In close collaboration with our industry partner GeneFluidics, we propose a rationale Product Development Plan for future clinical deployment.